Summary of Work: The "Mechanisms of Germ Cell Death" project aims to identify the mechanisms by which germ cells are killed in the testis. We earlier showed that the architecture of the normal seminiferous tubule was required for this apoptosis, which implicates a role for cell-cell adhesions and their related signals from the Sertoli cells. This had not been demonstrated previously. Working on glycol ether-induced apoptosis in pachytene spermatocytes, we showed that pharmaceuticals that inhibited calcium fluxes across membranes inhibited the apoptosis.Other labs have shown similar protection in thymus. However, recently, using fluorescent confocal laser microscopy, we demonstrated that such fluxes did not, in fact, occur, and we postulated alternative mechanisms, e.g., changes in membrane-bound kinases. Current studies have documented the recruitment of some PKC isoforms to around the dying cells, while other strategies are identifying the substrates that are phosphorylated during cell death, and which can be blocked when the lesion is also blocked. We plan to work backward to identify the kinases by first identifying the substrates. In the "Inhibited Spermiation" project, we have found N-cadherin in Sertoli cells, and desmoglein on late spermatids. Both these cadherins are in a distribution that suggests that they act to mediate spermatid adhesion. Importantly, we have found a suite of proteins inside the Sertoli cells that are known, in other cells, to regulate cadherin adhesiveness. Also, there is persistent cadherin staining around retained spermatids. Future immunoprecipitation studies should help identify protein-protein interactions that form the adhesions between Sertoli cells and spermatids, which will in turn lead to new understandings about how these adhesions are controlled, and interrupted by toxicants.